This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Multiple sclerosis (MS) affects approximately 400,000 Americans, two-thirds of whom are female. The causes of MS remain elusive, but a myelin protein is likely to be the autoantigen responsible for initiating the inflammatory response in the CNS, and sex steroids may modulate susceptibility to MS. Elucidation of the mechanisms by which these hormones influence microglial function and an understanding of the regulation of important myelin-specific proteins will be crucial in understanding the basis of gender differences in MS. The goals of this project are to determine molecular mechanisms that result in a gender bias in MS. Effects of female sex steroids upon the regulation of microglia function will be assessed by looking at expression levels and regulation of iNOS, a key enzyme in the production of inflammatory molecules responsible for the subsequent degradation of myelin characteristic of the disease. Transient transfections of iNOS-luciferase constructs into microglial cells will be done to examine this pathway, and an efficient system for these transfections has been optimized in the lab in recent months. Experiments are underway to assess iNOS expression in response to female sex steroids added to culture media. Modulation of the immune response by sex steroids is likely to contribute to the gender disparity of MS, but gender-specific expression of potential autoantigens in mediating susceptibility to the disease may also contribute. Existing data demonstrate expression of Plp mRNA encoding the most abundant protein found in mature CNS myelin, previously thought to be expressed exclusively in the CNS, in Leydig cells of the testes of mice, with no peripheral expression in female counterparts. We have obtained Western blot data confirming expression of Plp protein in testicular tissue. It is possible this peripheral expression could establish protective tolerance in males.